HistoClear II Protocol for Paraffin Removal and Tissue Processing
HistoClear II is a safer, xylene-free clearing agent used in histology for deparaffinization and tissue processing. Below is a general protocol for its use in histological applications:
1. Deparaffinization of Tissue Sections
Materials Needed:
HistoClear II Absolute ethanol (100%) Graded ethanol series (95%, 70%) Distilled water Glass slides with tissue sections Procedure:
Deparaffinization:
Immerse slides in HistoClear II for 2-3 changes, 5 minutes each. This step removes paraffin from tissue sections. Rehydration:
Transfer slides to 100% ethanol for 2 changes, 3-5 minutes each. Place slides in 95% ethanol for 2 minutes. Transfer to 70% ethanol for 2 minutes. Rinse in distilled water for 2 minutes. The tissue sections are now ready for staining. 2. Clearing Step in Histological Staining After dehydration in ethanol, HistoClear II is used as a clearing agent before mounting.
Procedure:
After dehydration in 100% ethanol, immerse slides in HistoClear II for 2-3 changes, 3-5 minutes each. Mount slides with a compatible mounting medium, such as HistoMount or another resin-based medium. 3. Tissue Processing for Embedding in Paraffin Materials Needed:
HistoClear II Absolute ethanol Paraffin wax Procedure:
Dehydration:
Immerse fixed tissue samples in 70% ethanol, 95% ethanol, and 100% ethanol, 1-2 hours each. Clearing with HistoClear II:
Immerse tissues in HistoClear II for 2 changes, 1 hour each. Paraffin Infiltration:
Transfer tissues to melted paraffin wax at 60°C for 2-3 changes, 1 hour each. Embedding:
Embed the tissue in paraffin blocks for sectioning.
